The present invention relates to compounds, compositions and methods using the same for preventing amyloid fibril formation. More particularly, the present invention relates to indole derivatives, compositions including same, and methods using the same, for the treatment of amyloid associated diseases, such as type II diabetes mellitus, Alzheimer's dementia or diseases, systemic and localized amyloidosis, and prion-related encephalopathies.
Amyloid material deposition (also referred to as amyloid plaque formation) is a central feature of a variety of unrelated pathological conditions including Alzheimer's disease, prion-related encephalopathies, type II diabetes mellitus, familial amyloidosis, secondary amyloidosis, and light-chain amyloidosis.
Amyloid material is composed of a dense network of rigid, nonbranching proteinaceous fibrils of indefinite length that are about 80 to 100 Å in diameter. Amyloid fibrils contain a core structure of polypeptide chains arranged in antiparallel β-pleated sheets lying with their long axes perpendicular to the long axis of the fibril [Both et al. (1997) Nature 385:787-93; Glenner (1980) N. Eng. J. Med. 302:1283-92].
Approximately twenty amyloid fibril proteins have been identified in-vivo and correlated with specific diseases. These proteins share little or no amino acid sequence homology, however the core structure of the amyloid fibrils is essentially the same. This common core structure of amyloid fibrils and the presence of common substances in amyloid deposits suggest that data characterizing a particular form of amyloid material may also be relevant to other forms of amyloid material and thus can be implemented in template design for the development of drugs for use against amyloid associated diseases such as type II diabetes mellitus, Alzheimer's dementia or diseases, and prion-related encephalopathies.
Furthermore, amyloid deposits do not appear to be inert in vivo, but rather are in a dynamic state of turnover and can even regress if the formation of fibrils is halted [Gillmore et al. (1997) Br. J. Haematol. 99:245-56].
Thus, therapies designed to inhibiting the production of amyloid polypeptides or inhibiting amyloidosis may be useful for treating amyloid associated diseases.
Inhibition of the production of amyloid polypeptides: Direct inhibition of the production of amyloid polypeptides may be accomplished, for example, through the use of antisense oligonucleotides, such as against human islet amyloid polypeptide messenger RNA (mRNA). In vitro, the addition of antisense oligonucleotides or the expression of antisense complementary DNA against islet amyloid polypeptide mRNA increases insulin mRNA and protein content of cells, demonstrating the potential effectiveness of this approach [Kulkarni et al. (1996) J. Endocrinol. 151:341-8; Novials et al. (1998) Pancreas 17:182-6]. However, no experimental result demonstrating the in vivo effectiveness of such antisense molecules has been disclosed.
Inhibition of the formation of amyloid fibrils: Amyloid, including islet amyloid, contains potential stabilizing or protective substances, such as serum amyloid P component, apolipoprotein E, and perlecan. Blocking their binding to developing amyloid fibrils could inhibit amyloidogenesis [Kahn et al. (1999) Diabetes 48:241-53], as could treatment with antibodies specific for certain parts of an amyloidogenic protein [Solomon et al. (1997) Proc. Natl. Acad. Sci. USA 94:4109-12].
The following summarizes current attempts to engineer drugs having the capability of destabilizing amyloid structures.
Destabilizing compounds: Heparin sulfate has been identified as a component of all amyloids and has also been implicated in the earliest stages of inflammation associated amyloid induction. Kisilevsky and co-workers (Mature Med. 1:143-148, 1995) described the use of low molecular weight anionic sulfonate or sulfate compounds that interfere with the interaction of heparin sulfate with the inflammation associated amyloid precursor and the β peptide of Alzheimer's disease (AD). Heparin sulfate specifically influences the soluble amyloid precursor (SAA2) to adopt an increased β-sheet structure characteristic of the protein folding pattern of amyloids. These anionic sulfonate or sulfate compounds were shown to inhibit heparin accelerated Aβ fibril formation, and were able to disassemble preformed fibrils in vitro, as monitored by electron micrography. Moreover, these compounds substantially arrested murine splenic inflammation associated amyloid progression in vivo in acute and chronic models. However, the most potent compound [i.e., poly-(vinylsulfonate)] showed acute toxicity. Similar toxicity has been observed with another compound, IDOX (Anthracycline 4′-iodo-4′-deoxy-doxorubicin), which has been observed to induce amyloid resorption in patients with immunoglobin light chain amyloidosis (AL) [Merlini et al. (1995) Proc. Natl. Acad. Sci. USA].
Destabilizing antibodies: Anti-β-amyloid monoclonal antibodies have been shown to be effective in disaggregating β-amyloid plaques and preventing β-amyloid plaque formation in vitro (U.S. Pat. No. 5,688,561). However, no experimental result demonstrating the in vivo effectiveness of such antibodies has been disclosed.
Small molecules: The potential use of small molecules which bind amyloid polypeptide and stabilize the native fold of the protein has been attempted in the case of transthyretin (TTR) protein [Peterson (1998) Proc. Natl. Acad. Sci. USA 95:12965-12960; Oza (1999) Bioorg. Med. Chem. Lett. 9:1-6]. Thus far, it has been demonstrated that molecules such as thyroxine and flufenamic acid are capable of preventing the conformation change, leading to amyloid formation. However, the use of such compounds in animal models has not yet been proven, and might be compromised due to the presence of such compounds in blood, or in proteins other than TTR, which are capable of binding these ligands.
Antioxidants: Another proposed therapy has been the intake of antioxidants in order to avoid oxidative stress and maintain amyloid proteins in their reduced state (i.e., monomers and dimers). The use of sulfite was shown to lead to more stable monomers of TTR protein both in vitro and in vivo [Altland (1999) Neurogenetics 2:183-188]. However, a complete characterization of the antioxidant effect is still not available and the interpretation of results concerning possible therapeutic strategies remains unclear.
Destabilizing peptides: The finding that the addition of synthetic peptides that disrupt the β-pleated sheets (‘β-sheet breakers’), thereby dissociating fibrils and preventing amyloidosis [Soto et al. (1998) Nat. Med. 4:822-6,], is particularly promising from a clinical point of view. In that disclosure, it was found that a penta-residue peptide inhibited amyloid beta-protein fibrillogenesis, disassembled preformed fibrils in vitro, and prevented neuronal death induced by fibrils in cell culture. In addition, the beta-sheet breaker peptide significantly reduced amyloid beta-protein deposition in vivo, and completely blocked the formation of amyloid fibrils in a rat brain model of amyloidosis.
Green tea extracts: U.S. Patent Applications having the Publication Nos. 20020086067 and 20020151506 teach the use of various components of green tea extracts for treating an amyloid disease. While these patent applications teach that these components inhibit amyloid fibril formation, they fail to teach neither a mechanism nor a common structural feature which provides these green tea components with such an activity. The present inventor has previously shown that aromatic interactions play a key role in amyloid fibril formation by serving as structural and functional elements that direct molecular recognition and self-assembly and have further shown that such aromatic interactions can be induced by tryptophane containing peptide sequences [Gazit (2002) FASEB J. 16:77-83].
As is well known, tryptophane is an amino acid residue having an indole side-chain. Thus, while conceiving the present invention, it was envisioned that compounds having an aromatic moiety and particularly an indole moiety, which may participate in such aromatic interactions, could efficiently serve as inhibitors of amyloid fibril formation. It was further envisioned that since indoles are known to act as antioxidants [Herraiz et al. (2004) Free Radic Res. 38:323-31], compounds having an indole moiety could serve simultaneously as inhibitors of amyloid fibril formation and as antioxidants and as such may exert synergistic effect in treating amyloid-associated diseases.